A defect in the presentation of intracellular viral antigens is restored by interferon-gamma in cell lines with impaired major histocompatibility complex class I assembly.

Surface expression of the majority of class I major histocompatibility complex (MHC) heavy chains is known to require assembly with beta 2 microglobulin (beta 2m). To define other factors involved in class I MHC assembly, we have studied two tumor cell lines that are deficient in cell surface class I (H-2) expression. The BC2 fibrosarcoma and the CMT lung carcinoma express only intracellular unassociated heavy chains despite the presence of beta 2m. As described previously, when these cell lines are treated with interferon-gamma (IFN-gamma), they are capable of assembling and transporting class I molecules to the cell surface. In this study, we have shown that in the absence of IFN-gamma these mutant cells are unable to present intracellular viral antigens, although they can be lysed by specific cytotoxic T lymphocyte (CTL) after pre-incubation with the corresponding synthetic peptide. Flow cytometric analysis demonstrated that extracellular peptide was capable of increasing twofold the surface expression of beta 2m-heavy chain complexes. Furthermore, immunoprecipitation experiments confirmed that peptide stabilizes chain association in the BC2 cell lysates. However, infecting these mutants with vectors expressing either pre-processed antigen or rapidly degraded antigen, failed to overcome their defect in the presentation of endogenous peptide to specific CTL or to mediate surface expression of class I MHC. Preincubation with IFN-gamma completely reversed the endogenous peptide presentation defect, even in mutant cells transfected with a vector encoding a cDNA for the H-2 molecule restricting CTL recognition. This last result suggests that IFN-gamma corrects the defect by a mechanism separate from simple enhancement of the number of class I molecules produced by the cell. Because there is growing evidence that endogenous peptides can participate in class I MHC assembly, the defect in these mutants could be ascribed to the lack of access to class I molecules by the endogenous peptide. This would prevent stable association of the heavy and light chains and their subsequent transport. Our data suggests that IFN-gamma reestablishes class I MHC surface expression by restoring access of endogenously synthesized peptide to class I molecules.     

Differential effect of transporter Tap 2 gene introduction into RMA-S cells on viral antigen processing

The protein products of the Tap (Transporter associated with antigen processing) 1 and 2 genes are presumed to deliver peptides across the endoplasmic reticulum (ER) for assembly with major histocompatibility complex (MHC) class I molecules. The antigen processing-defective cell line RMA-S (H-2^b) has a premature stop in the Tap 2 gene and probably therefore fails to deliver peptides into the ER, which leads to a low level of cell surface MHC class I molecules. Transfection of a Tap 2 gene restores to RMA-S both MHC class I molecule expression and the ability to present influenza viral antigens. We investigated the ability of RMA-S cells transfected with a Tap 2 gene to process and present alloantigens, Sendai and Rauscher viral antigens to allogeneic and virus-specific cytotoxic T lymphocytes. We found that allogeneic peptides as well as Rauscher and Sendai viral peptides can be processed and presented by RMA-S but at reduced levels. Transfection of a Tap 2 gene of mouse (BALB/c, H-2^d) or rat origin into RMA-S increased the presentation of Sendai viral antigens and partially restored the presentation of allogeneic antigens. The already low level of Rauscher viral peptides presented by RMA-S is not elevated by transfection of either Tap 2 gene into RMA-S. This indicates a differential effect of transfection of a Tap 2 gene of rat or allogeneic mouse origin into RMA-S on viral antigen processing.     

Cytotoxic T lymphocytes against the antigen-processing-defective RMA-S tumor cell line.

RMA-S is an antigen processing-defective cell line, obtained from a Rauscher virus-induced tumor. The cells express only a low level of cell surface major histocompatibility complex (MHC) class I molecules, which are supposed to be devoid of internally derived antigenic peptides. We investigated Rauscher virus expression and Rauscher peptide presentation to virus-specific cytotoxic T lymphocytes (CTL) by this cell line. Viral proteins are expressed properly, both intracellularly and at the cell surface of RMA-S. Rauscher peptides are presented to virus-specific CTL in the groove of both the class I H-2Kb and Db molecules, but at a low level. Culture of RMA-S cells at room temperature increases their susceptibility to CTL. The RMA-S defect thus affects, but not totally abrogates, Rauscher peptide presentation by MHC class I molecules via the endogenous pathway. This indicates that the RMA-S antigen processing deficit is not absolute.     

Peptide loading of empty major histocompatibility complex molecules on RMA-S cells allows the induction of primary cytotoxic T lymphocyte responses.

The antigen processing-defective mutant cell line RMA-S expresses at the cell surface major histocompatibility complex (MHC) class I molecules devoid of peptide that can be efficiently loaded with exogenous immunogenic peptides. We now report that viral peptide-loaded RMA-S cells, unlike parental RMA cells, can induce primary cytotoxic T lymphocyte (CTL) responses in vitro, in a T helper cell-independent fashion. This was shown for an H-2Kb-binding peptide of Sendai virus nucleoprotein and an H-2Db-binding peptide of adenovirus type 5 E1A protein with responding spleen cells of C57BL/6 mice, the strain of origin of RMA and RMA-S cells. Primary Sendai peptide-induced CTL lyse both peptide-loaded and virus-infected cells. Pre-culture of RMA-S cells at low temperature (22 degrees - 26 degrees C), which increases the amount of empty MHC class I molecules at the cell surface, decreases the peptide concentrations required for the induction of primary CTL responses. Primary peptide-specific CTL responses induced by peptide-loaded RMA-S cells are CD4+ cell- and MHC class II+ cell-independent. CTL response induction is blocked by the presence of anti-CD8 monoclonal antibody during culture. Direct peptide binding studies confirm the efficient loading of empty MHC molecules on RMA-S cells with peptide and show 2.5-fold more peptide bound per RMA-S cell compared to RMA cells. An additional factor explaining the difference in primary response induction between RMA and RMA-S cells is related to the CD8 dependence of these responses. MHC class I molecules occupied with irrelevant peptides (a majority present on RMA, largely absent on RMA-S) may interfere in the interaction of the CD8 molecule with relevant MHC/peptide complexes. The results delineate a novel strategy of peptide based in vitro immunization to elicit CD8+ cytotoxic T cell responses.     

Different types of allospecific CTL clones identified by their ability to recognize peptide loading-defective target cells

Allospecific immune responses against the MHC of another individual are remarkably strong, due t a high number of responding T cell clones. Although it has been demonstrated that some allospecific cytotoxic T lymphocytes (CTL) recognize peptides presented by allogeneic MHC class I molecules, it has remained unclear whether MHC molecules can be recognized directly. We used the H-2b-derived murine lymphoma mutant RMA-S, which has a defect affecting peptide loading of class I molecules, to test whether recognition by allospecific CTL always requires the presence of peptides. Three types of anti-H-2Kb CTL clones can be distinguished by their ability to lyse RMA-S target cells. Type A CTL clones efficiently lyse these target cells, the lysis by type B CTL clones is inefficient, and type C clones fail to lyse RMA-S. Up-regulation of the levels of H-2Kb density improved lysis by type B clones, but did not lead to lysis by type C clones. Some type A and B CTL clones apparently can recognize class I molecules devoid of peptides, while others are likely to recognize peptides which are not affected by the presentation defect of RMA-S. We suggest that type C clones are specific for peptides which are not presented by the mutant cells. The results show that the majority of alloreactive CTL recognize peptide/MHC complexes, while some CTL behave as if they can recognize class I molecules in the absence of MHC-bound peptides.     

TAP2-defective RMA-S cells present Sendai virus antigen to cytotoxic T lymphocytes

The murine antigen-processing-defective mutant cell line RMA-S is leaky in the presentation of certain endogenously synthesized minor histocompatibility and viral antigens to major histocompatibility complex (MHC) class I-restricted cytotoxic T lymphocytes (CTL). The viral antigens include influenza virus nucleoprotein, vesicular stomatitis virus (VSV) nucleocapsid and Rauscher murine leukemia virus (MuLV) antigen. Here we demonstrate Sendai virus antigen presentation by the HAM2 (murine TAP2, transporter associated with antigen presentation type 2)-defective RMA-S cell line and compare antigen presentation after restoration of the defect by murine TAP1/2 gene transfection. Kinetic studies revealed that RMA-S cells required 2-3 h longer incubation and approximately 10 times higher doses of Sendai virus to reach the same level of killing as the RMA parental line. After transfection of RMA-S cells with the murine TAP1/2 gene, Sendai virus antigen presentation was restored to levels of the RMA wild-type line with regard to time of virus infection and dose of virus needed for sensitizing target cells. The presentation of Sendai virus antigen in RMA-S cells was sensitive to brefeldin A (BFA), suggesting that the presentation was mediated via the endogenous pathway. Our findings comfirmed leakiness of antigen presentation in RMA-S cells and extended it to Sendai virus. The results underscored the role for intact expression of the TAP 1/2 molecules for efficient MHC class I-mediated antigen presentation.     

Surface expression of β2-microglobulin-associated thymus-leukemia antigen is independent of TAP2

Mouse thymus-leukemia antigen (TL), like other major histocompatibility complex (MHC) class I-b antigens, displays signs of a specialized function. It is normally expressed at high levels on immature thymocytes and at moderate levels on gut epithelium and activated mature T cells. A promoter/enhancer region unique among class I genes accounts for this narrow range of tissue distribution. Like most other class I molecules, TL is dependent upon endogenous β2-microglobulin (β2m) for transport to the surface. However, here we show that unlike most other MHC class I molecules, TL is expressed efficiently in the absence of functional transporter associated with antigen processing subunit 2 (TAP2). A putative fourth TL^a gene cloned from A.SL1 cells was expressed in RMA and RMA-S cells. In bulk transformants, TL expression is higher in TAP2^? RMA-S cells than in wild-type RMA cells, and is not elevated by incubation at reduced temperatures or exposure to exogenous β2m. Analysis of immunoprecipitasted molecules by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate indicates that TL is processed normally in RMA-S cells and is associated with β2m both intracellularly and at the cell surface. However, TL heavy chains expressed on the cell surface in the absence of TAP2 are cleaved to a predominant 38 kDa fragment, presumably the result of an altered conformation that renders TL more susceptible to proteolysis. These results suggest that while TL may normally acquire TAP2-dependent peptides, this class I-b molecule does not require them for efficient export to, and stable expression at the cell surface.     

Macrophages present exogenous antigens by class I major histocompatibility complex molecules via a secretory pathway as a consequence of interferon-gamma activation

Macrophages can process and present exogenous antigens on major histocompatibility complex (MHC) class I molecules through an alternative mechanism involving the internalization of antigens and the secretion of peptides loading MHC class I molecules at the cell surface. In this paper, we found that interferon-gamma (IFN-gamma) -activated macrophages infected with Salmonella typhimurum secreted peptides able to load empty MHC Kb molecules on co-cultured TAP-2-deficient RMA-S cells, added as targets for peptide loading. The increase in class I Kb on the RMA-S cells, resulting from the macrophage-derived peptides, exhibited a comparable stability as the direct addition of an exogenous Kb-binding peptide (OVA257-264) to the RMA-S cells. In both cases, the Kb complexes were stable for at least 3 hr after separating the RMA-S cells from the macrophages. The endosomal inhibitors, leupeptin and ammonium chloride, did not inhibit the release of peptides and the increase in Kb staining on the RMA-S cells in the co-culture systems. Brefeldin A also had no effect. P815 cells previously co-cultured with Salmonella-infected macrophages became targets for cytotoxic T lymphocytes isolated from Salmonella-infected BALB/c mice. Taken together, our data suggest that IFN-gamma-activated macrophages process exogenous antigens in an intracellular compartment where serine proteases generate peptides released to the external environment for loading empty MHC class I molecules at the cell surface. This TAP-independent mechanism for the MHC class I presentation may be involved in priming cytotoxic T lymphocytes against intracellular pathogens in vivo.     

Intradomain H-2Kd/Dd recombinants define the same regions as crucial for recognition by alloreactive or major histocompatibility complex-restricted cytolytic T cells

To identify residues on class I major histocompatibility complex (MHC) antigens that are important in T cell recognition, we analyzed a series of 11 intradomain recombinant mouse MHC (H-2) molecules in which N-terminal H-2Kd segments of varying lengths are followed by H-2Dd segments. Lysis of L cell transfectant target cells by a series of alloreactive cytolytic T cell (CTL) clones specific for Kd or for Dd revealed several regions that contain residues critical for specific recognition. These residues map within the presumed antigen-binding site of the MHC molecule. Of particular interest was the finding that the two regions identified as important for Kd allorecognition match those that influence the recognition of synthetic peptide antigens by Kd-restricted CTL.     

Presentation of viral antigens restricted by H-2Kb,Db or Kd in proteasome subunit LMP2-and LMP7-deficient cells

In the class II region of the major histocompatibility complex (MHC), four genes implicated in MHC class I-mediated antigen processing have been described. Two genes (TAP 1 and TAP 2) code for multimembrane-spanning ATP-binding transporter proteins and two genes (LMP 2 and LMP 7) code for subunits of the proteasome. While TAP 1 and TAP 2 have been shown to transport antigenic peptides from the cytosol into the endoplasmic reticulum, where the peptides associate with MHC class I molecules, the role of LMP 2/7 in antigen presentation is less clear. Using antigen processing mutant T2 cells that lack TAP 1/2 and LMP 2/7 genes, it was recently shown that expression of TAP 1/2 alone was sufficient for processing and presentation of the influenza matrix protein M1 as well as the minor histocompatibility antigen HA-2 by HLA-A2. To understand if presentation of a broader range of viral antigens occurs in the absence of LMP 2/7, we transfected T2 cells with TAP 1, TAP 2 and either of the H-2K^b, D^b or K^d genes and tested their ability to present vesicular stomatitis vires and influenza virus antigens to virus-specific cytotoxic T lymphocytes. We found that T2 cells, expressing TAP 1/2 gene products, presented all tested viral antigens restricted through either the H-2K^b, D^b or K^d class I molecules. We conclude that the proteasome subunits LMP 2/7 as well as other gene products in the MHC class II region, except from TAP 1/2, are not generally necessary for presentation of a broader panel of viral antigens to cytotoxic T cells. However, the present results do not exclude that LMP 2/7 in a more subtle way may, or in rare cases completely, affect processing of antigen for presentation by MHC class I molecules.     

Expression of medial class I histocompatibility antigens on RMA-S mutant cells

The RMA-S mutant T cell line is defective in H-2b restricted antigen presentation and has markedly reduced surface expression of Kb and Db. We examined RMA-S for the expression of the medial class I histocompatibility antigens Qa1b and Mta. While RMA-S targets varied in their susceptibility to lysis by cytotoxic T lymphocytes (CTL) specific for Qa1b, Mta levels were detectable but consistently low compared to the parent RMA cell line. Addition of synthetic ND1 alpha 1-26 or ND1 alpha 1-17 peptides that mimic MTF alpha (the ligand of Mta) increased killing of RMA-S by anti-Mta alpha CTL to levels comparable to or better than RMA, with 300 nM peptide being fully effective. None of the MTF peptides increased the killing of RMA-S by anti-H-2b or anti-Qa1b CTL, even at the highest (1 microM) peptide concentrations. RMA-S cells treated with 100 microM of either the ND1 alpha 4-26 or ND1 alpha 1-26 peptides showed a small increase in the fluorescent staining for beta 2-microglobulin but not for H-2Kb or H-2Db. These results show that Mta and Qa1b, although affected, are not obliterated by the defect in RMA-S cells; that the association of MTF peptides with HMT is exclusive; and that MTF enters the endoplasmic reticulum in the same fashion as other endogenous peptides.     

Localization of the conformational alteration of MHC molecules induced by the association of mouse class I heavy chain with a xenogeneic beta 2-microglobulin.

Changes in the antigenicity of major histocompatibility complex (MHC) class I molecules resulting from the association of bovine beta 2-microglobulin (beta 2-m) with mouse class I heavy chains were investigated. Mice (H-2b) were immunized with syngeneic Concanavalin A (Con A) blasts induced in the presence of fetal calf serum (FCS) in conditions allowing exchange between mouse and bovine beta 2-microglobulin (beta 2-m). Spleen cells from hyperimmunized mice were fused with myeloma cells and two monoclonal antibodies which required for their reactivity the presence of FCS have been further studied. One of them (CAB 297) recognized a determinant of bovine beta 2-m which is present on free molecules in solution as well as when they are associated with either mouse or bovine class I heavy chains. In contrast, the second monoclonal antibody (CBB 70) did not react with free bovine beta 2-m molecules, nor with beta 2-m associated with bovine class I heavy chains. It did react with cells of some H-2 haplotypes (b, f, p and r) but only when their class I heavy chains are associated with bovine or with human beta 2-m. Therefore, expression of the CBB 70 defined antigenic determinant requires both xenogeneic beta 2-m and class I heavy chain of a given H-2 molecule. In order to precisely localize the antigenic determinant defined by this monoclonal antibody and therefore the region altered by the association of class I heavy chain with xenogeneic beta 2-m, we made use of exon shuffled class I molecules. The results indicate that changes induced by the association of bovine beta 2-m with H-2 class I heavy chain affect the conformation of the alpha 2 domain. These studies illustrate that MHC class I molecules exhibit a considerable conformational flexibility which could influence their ability to bind and present various peptides to the T-cell receptor.     

The conformational flexibility of class I H-2 molecules as revealed by anti-peptide antibodies specific for intracytoplasmic determinants: differential reactivity of beta 2-microglobulin "bound" and "free" H-2Kb heavy chains.

Immunoprecipitation experiments using anti-peptide antisera prepared against exon 6, exon 7 and exon 8-encoded intracytoplasmic regions of the H-2Kb gene product indicated that approximately 1/3 of the H-2Kb heavy chains in a cell surface-labelled glycoprotein fraction from EL-4 cells, or H-2b spleen cells, is not associated with beta 2-microglobulin (beta 2-m). This population of "free" H-2Kb heavy chains failed to react with alloantisera or monoclonal antibodies specific for conventional H-2Kb serological determinants, suggesting that significant conformational alterations were induced in the extracellular domains upon dissociation of beta 2-m. In addition, although antibodies to intracytoplasmic peptide 8 were able to react with both "free" and beta 2-m "bound" heavy chains, the determinants seen by anti-peptide 6 and anti-peptide 7 were only recognized in the "free" heavy chain. These data suggest that the conformational perturbation of the extracellular domains induced by beta 2-m dissociation can be "transmitted" to the intracytoplasmic region of the heavy chain. These results indicate the potential for a class I heavy chain-mediated transmembrane signalling event, and suggest that the "free" class I heavy chain might have a role to play in the major histocompatibility complex (MHC)-restricted presentation of T-cell determinants to cytotoxic T-lymphocytes.     

Identification of murine H-2Db histocompatibility antigens in cells transfected with cloned H-2 genes

Clones of mouse L-cells transformed with 21 cosmids containing 15 major histocompatibility complex class I genes of C57BL10 (H-2b) sperm cell DNA were analyzed for the expression of their transfected H-2 and Qa/Tla genes. Three cosmids contained a single gene, mapping to the H-2D region. This gene encodes the H-2Db alloantigen: mouse L-cells transfected with cosmids containing this gene reacted with monoclonal antibodies and alloantisera specific for the H-2Db antigen and expressed a 46-kd H-2 heavy chain associated with beta 2-microglobulin in their cell membranes. Furthermore, these transfected cells were stimulators of, and targets for, anti-H-2Db cytotoxic T-lymphocytes. Eighteen cosmids contained 14 different genes mapping to the Qa and Tla regions. L-cells transfected with these genes did not express class I genes reacting with alloantisera or monoclonal antibodies against Qa2, Qa4 or TL differentiation antigens. In particular, the Qa2,3 gene of C57BL10 was not identified.     

Influence of glycosylphosphatidylinositol-linked H-2Dd molecules on target cell protection and natural killer cell specificity in transgenic mice

The expression of certain major histocompatibility complex (MHC) class I ligands on target cells is one important determinate of their susceptibility to lysis by natural killer (NK) cells. NK cells express receptor molecules that bind to MHC class I. Upon binding to their MHC class I ligand, the NK cell is presumed to receive a signal through its receptor that inhibits lysis. It is unclear what role the MHC class I molecules of the effector and target cells play in signaling to the NK cell. We have investigated the role of the cytoplasmic and transmembrane domains of MHC class I molecules by producing a glycosylphosphatidylinositol (GPI)-linked H-2D^d molecule. The GPI-linked H-2D^d molecule is recognized by H-2D^d-specific antibodies and cytotoxic T lymphocytes. Expression of the GPI-linked H-2D^d molecule on H-2^b tumor cells resulted in protection of the tumor cells after transplantation into D8 mice (H-2^b, H-2D^d) from rejection by NK cells. In addition, NK cells from mice expressing the GPI-linked H-2D^d molecule as a transgene were able to kill nontransgenic H-2^b lymphoblast target cells. The GPI-linked MHC class I molecule was able to alter NK cell specificity at the target and effector cell levels. Thus, the expression of the cytoplasmic and transmembrane domains of MHC class I molecules are not necessary for protection and alteration of NK cell specificity.     

Binding of divalent H-2Kd/IgG2aFc fusion protein to murine macrophage via Fc-FcR interaction

Peptide-MHC class I complex （pMHC） is a specific ligand for TCR recognition, and important for CD8^＋T cell activation. Here we described a genetically engineered divalent class I major histocompatibility complex （MHC） molecule, H-2K^d/IgG2aFc, a fusion protein consisting of the extracellular domains of H-2K^d, a murine MHC class I molecule, and the Fc region of IgG2a. This fusion protein is expected to attach the H-2K^d molecule to the surface of murine macrophage （MФ） through its Fc portion binding to Fc receptor （FcR） of MФ. cDNAs coding for the extracellular domains of H-2K^d and the Fc region of IgG2a were cloned respectively, and then recombined into plasmid pcDNA3.1（＋）. The H-2K^d/IgG2aFc protein was expressed by the plasmid-transfected cell line J558L, and purified from its supernatant with a Staphylococcal Protein A （SPA） column. The fusion protein showed a 58.4 kDa band as revealed by SDS-PAGE and Western blotting with murine IgG-specific antibody, which consists with that expected for extracellular domains of H-2K^d heavy chain plus the Fc region of IgG2a. The sandwich ELISA assay with antibodies specific for Fc portion and for H-2K^d indicated the fusion protein consists of both Fc portion and H-2K^d. Peritoneal MФ of C57BL/6 （H-2K^b） can be stained with H-2K^d specific monoclonal antibody （mAb） after incubated with the H-2K^d/IgG2aFc fusion protein. These results demonstrate the fusion protein can be used to attach the H-2K^d molecule to the surface of murine MФ, and provides a novel means to manipulate the T cell recognized epitope on the surface of murine MФ, which can be applied to activate antigen-specific cytotoxic T lymphocyte （CTL）.     

Differential association of beta2-microglobulin mutants with MHC class I heavy chains and structural analysis demonstrate allele-specific interactions.

Dynamic interactions between major histocompatibility complex (MHC) class I heavy chains and beta2-microglobulin (beta2m) play a critical role in their stability on the cell surface, and their ability to present peptide antigens to CD8+ T-cells. A cursory review of protein sequence homologies and three-dimensional crystal structures of MHC complexes might indicate very similar modes of interaction between the heavy and light chains. In this report, a panel of human beta2m mutants was screened to probe the interactions of beta2m with the murine MHC molecules H-2Kb, -Db, -Kd, -Ld, and -Dd. Binding experiments coupled with analyses of existing three-dimensional crystal structures demonstrate allelic differences in their interaction with beta2m. A comprehensive analysis of the existing murine MHC structures indicates a conformational flexibility on the part of murine beta2m that is not present in beta2m of the human structures. This flexibility is in a region directly interacting with the heavy chain and may relate to its lower affinity for murine heavy chains relative to human beta2m. This defined panel of beta2m mutants of differing affinity may also be useful for subsequent studies of thymic selection, T-cell recognition, and more refined algorithms for protein structure prediction.     

A novel H-2s class I molecule expressed on a B-cell leukemia from SJL/J mice

Anti-H-2.33 [(B10.D2 X A)F1 anti-B10.A(5R)], which predominantly contains antibodies recognizing H-2Kb and IAb molecules, was found to be cytotoxic against DMLM 1678, a B-cell leukemia of SJL/J (H-2s) origin. The antiserum precipitated a typical class I (H-2-like) molecule from labeled tumor cell preparations as judged by molecular mass, papain susceptibility and association with beta 2-microglobulin. Sequential immunoprecipitation studies revealed that it was distinct from either H-2Ks or H-2Ds, the 2 molecules expressing the private antigens of the H-2s haplotype. Absorption analysis using congenic mice mapped the gene controlling the expression of the novel molecule telomeric to the S-region within the major histocompatibility complex.     

Isolation and characterization of mouse beta 2-microglobulin allotypes

Two allelic forms of mouse beta 2-microglobulin (beta 2m), the smaller polypeptide chain of the H-2 histocompatibility antigens, were purified from urine and partially characterized. The isoelectric points of the two allotypes are 7.4 (beta 2ma) and 8.1 (beta 2mb). The electrophoretic mobility of beta 2ma is decreased by reduction, whereas the mobility of beta 2mb is not. Urinary beta 2m can replace endogenous beta 2m in mouse H-2 histocompatibility antigens.